Journal: Nature Communications

Article Title: Spatiotemporal interplay between epithelial and mesenchymal cells drives human dentinogenesis

doi: 10.1038/s41467-026-69545-3

Figure Lengend Snippet: a Split violin plots showing the expression distribution of WNT and NOTCH pathway components in DE (red) and DPL (blue). Each dot represents a single cell. Adjacent pie charts quantify the percentage of cells within the DE population expressing specific WNT and NOTCH ligands. b Dot plots illustrating the dynamic expression of WNT and NOTCH pathway genes in DE (left panel, red color scale) and DPL (right panel, blue color scale) across developmental stages. Dot size is proportional to the percentage of cells expressing the gene, and color intensity reflects the average z-score normalized expression level. PCW, post-conception weeks; Y, years. c Line graphs tracking the average expression of key WNT ( WNT6 , WNT7B , WNT10B ) and NOTCH ( JAG1 ) ligands within the DE population over time. d IF validation of the spatial localization of key WNT and NOTCH pathway proteins across different developmental stages. Target proteins (WNT6, WNT7B, WNT10B, DKK1, SFRP1, LRP6, LEF1, JAG1, NOTCH2, HEY1) are shown in green or red, with nuclei counterstained with DAPI (blue). Dotted lines outline the epithelial-mesenchymal boundary. Insets show magnified views of the indicated regions. Images are representative of experiments that were independently repeated three times with similar results ( n = 3 biological replicates). PT permanent tooth. Scale bars: 100 μm. Data in ( a – c ) are derived from scRNA-seq analysis of n = 2 biological replicates per developmental stage.

Article Snippet: Primary antibodies were applied against: DSPP (Abcam, ab216892), Ki67 (Cell Signaling Technology, 9129, Clone D3B5), KRT14 (Abcam, ab119695, Clone LL002), WNT6 (Abcam, ab50030), WNT7B (Proteintech, 10605-1-AP), WNT10B (Abcam, ab70816), LRP6 (Santa Cruz, sc-25317, Clone C-10), JAG1 (Abcam, ab300561, Clone EPR26388-51), NOTCH2 (Abcam, ab313453, Clone EPR28701-38), DKK1 (Abcam, ab61034, Clone EPR4759), SFRP1 (Proteintech, 26460-1-AP), HEY1 (Proteintech, 19929-1-AP), LEF1 (Abcam, ab137872, Clone EPR2029Y), BMP2 (Abcam,ab214821), BMP4 (Abcam, ab39973), GFP(CST, 2956, Clone D5.1) and rabbit IgG isotype control (CST, 3900, Clone DA1E) were added.

Techniques: Expressing, Single Cell, Biomarker Discovery, Derivative Assay

Journal: Aging (Albany NY)

Article Title: Hydrogen sulfide is a novel regulator implicated in glucocorticoids-inhibited bone formation

doi: 10.18632/aging.102269

Figure Lengend Snippet: Involvement of the Wnt/β-catenin pathway in the protective effect of GYY4137 against Dex-induced osteoblast activities. ( A ) mRNA expression of Wnt-signaling target genes was measured in primary osteoblast pretreated with Dex or/and GYY4137. n=3, *p<0.05, **p<0.01. ( B – C ) Immunofluorescence staining of the osteoblast after a 2-day incubation with Dex or/and GYY4137. ( D – E ) Effect of the wnt/β-catenin inhibitor on the protein expression of wnt3a, wnt6, β-catenin at 48hr with GYY4137 pretreatment in Dex-induced osteoblast activities. n=3, *p<0.05, **p<0.01.

Article Snippet: Once the osteoblasts reached 50–70% confluence, the cells were infected with Wnt3a or Wnt6 shRNA lentiviral particles (Santa Cruz Biotechnology), respectively.

Techniques: Expressing, Immunofluorescence, Staining, Incubation

Journal: Aging (Albany NY)

Article Title: Hydrogen sulfide is a novel regulator implicated in glucocorticoids-inhibited bone formation

doi: 10.18632/aging.102269

Figure Lengend Snippet: Knockdown of Wnt3a or Wnt6 abolished the GYY4137 effect on Dex-treated osteoblast proliferation and differentiation. ( A ) All cells expressed GFP, showing that the cells were infected by lentivirus. ( B – C ) Verified Wnt3a or Wnt6 knockdown effect by lentivirus-mediated transduction of primary culture osteoclasts precursors. n=3, **p<0.01. ( D ) The proliferation of rat primary osteoblasts cells was measured by CCK8 assay after the cells were infected by lentivirus. ( E ) Representative photomicrographs (x200) of EdU staining and corresponding total cell photomicrographs. Blue: Hoechst labeling of cell nuclei; red: EdU labeling of nuclei of proliferative cells. ( F ) Quantitative data showing the percentages of EdU-positive cells in different treatment groups (number of red versus number of blue nuclei). n=3, **p<0.01. ( G ) Western blot analysis of Runx2 and Osterix expression in rat primary osteoblasts of each group. n=3, **p<0.01. ( H ) ALP staining was measured in primary osteoblast per group.

Article Snippet: Once the osteoblasts reached 50–70% confluence, the cells were infected with Wnt3a or Wnt6 shRNA lentiviral particles (Santa Cruz Biotechnology), respectively.

Techniques: Knockdown, Infection, Transduction, CCK-8 Assay, Staining, Labeling, Western Blot, Expressing

Journal: Phytomedicine : international journal of phytotherapy and phytopharmacology

Article Title: Magnolol regulates miR-200c-3p to inhibit epithelial-mesenchymal transition and retinoblastoma progression by modulating the ZEB1/E-cadherin axis in vitro and in vivo.

doi: 10.1016/j.phymed.2022.154597

Figure Lengend Snippet: Fig. 2. MAG regulates the Wnt16/β-catenin/ZEB1/E-cadherin signaling pathway to inhibit EMT in Y79 cells. (a) Y79 cells were treated with various doses of MAG for 24 h. mRNA expression of Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA was examined through RT-qPCR analysis. **p < 0.01, ***p < 0.001, ****p < 0.0001 compared with the control group. (b–c) Protein expression of Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA was examined via immunofluorescent staining and western blotting. Scale bar of confocal microscope image = 4 μm. Cytosolic (CP) and nuclear protein (NP). Each group: n = 3.

Article Snippet: Briefly, the cells were cultured on glass coverslips, washed with cold phosphate-buffered saline (PBS), and fixed with 4% paraformaldehyde in PBS at 4 ◦C for 15 min. After blocking, the cells were incubated with primary antibodies against Wnt16 (Santa Cruz), Y.-H. Lai et al. Phytomedicine 110 (2023) 154597 β-catenin (Gene Tex), ZEB1 (Novusbio), E-cadherin (Cell Signaling Technology), FN1 (Gene Tex), and VEGFA (Santa Cruz) overnight at 4 ◦C, rinsed with PBS, and incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature, followed by staining with 4’,6-diamidino-2-phenylindole (DAPI; Invitrogen).

Techniques: Expressing, Quantitative RT-PCR, Control, Staining, Western Blot, Microscopy

Journal: Phytomedicine : international journal of phytotherapy and phytopharmacology

Article Title: Magnolol regulates miR-200c-3p to inhibit epithelial-mesenchymal transition and retinoblastoma progression by modulating the ZEB1/E-cadherin axis in vitro and in vivo.

doi: 10.1016/j.phymed.2022.154597

Figure Lengend Snippet: Fig. 5. MAG attenuates the Wnt16/β-catenin/ZEB1 signaling pathway and EMT in vivo. (a) Expression of Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA mRNA in subcutaneous xenograft model tumors were examined through RT-qPCR. ***p < 0.001, ****p < 0.0001 compared with the control group. Protein expression of Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA in (b) subcutaneous xenograft model tumors, (c) OX animal model, and (d) inguinal lymph nodes was examined through immunohistochemistry staining. Scale bar = 20 µm. Each group: n = 6.

Article Snippet: Briefly, the cells were cultured on glass coverslips, washed with cold phosphate-buffered saline (PBS), and fixed with 4% paraformaldehyde in PBS at 4 ◦C for 15 min. After blocking, the cells were incubated with primary antibodies against Wnt16 (Santa Cruz), Y.-H. Lai et al. Phytomedicine 110 (2023) 154597 β-catenin (Gene Tex), ZEB1 (Novusbio), E-cadherin (Cell Signaling Technology), FN1 (Gene Tex), and VEGFA (Santa Cruz) overnight at 4 ◦C, rinsed with PBS, and incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature, followed by staining with 4’,6-diamidino-2-phenylindole (DAPI; Invitrogen).

Techniques: In Vivo, Expressing, Quantitative RT-PCR, Control, Animal Model, Immunohistochemistry, Staining

Journal: Phytomedicine : international journal of phytotherapy and phytopharmacology

Article Title: Magnolol regulates miR-200c-3p to inhibit epithelial-mesenchymal transition and retinoblastoma progression by modulating the ZEB1/E-cadherin axis in vitro and in vivo.

doi: 10.1016/j.phymed.2022.154597

Figure Lengend Snippet: Fig. 7. miR-200c-3p suppresses Wnt16/β-catenin/ZEB1 pathway activation and attenuates EMT-associated protein expression in vitro. Y79 cells were transfected with miR-200c-3p mimic or inhibitor for 48 h. (a) miR-200c-3p expression was examined through RT-qPCR. ****p < 0.0001 compared with the control group. (b) mRNA expression of Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA was detected through RT-qPCR. Ns, no significance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 compared with the control group. (c–d) Protein expression of Wnt16, β-catenin, ZEB1, E-cadherin, FN1, α-SMA, and VEGFA was examined through immunocytochemistry staining and western blotting. Scale bar of confocal microscope image = 4 µm. Cytosolic (CP) and nuclear protein (NP). Each group: n = 3.

Article Snippet: Briefly, the cells were cultured on glass coverslips, washed with cold phosphate-buffered saline (PBS), and fixed with 4% paraformaldehyde in PBS at 4 ◦C for 15 min. After blocking, the cells were incubated with primary antibodies against Wnt16 (Santa Cruz), Y.-H. Lai et al. Phytomedicine 110 (2023) 154597 β-catenin (Gene Tex), ZEB1 (Novusbio), E-cadherin (Cell Signaling Technology), FN1 (Gene Tex), and VEGFA (Santa Cruz) overnight at 4 ◦C, rinsed with PBS, and incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature, followed by staining with 4’,6-diamidino-2-phenylindole (DAPI; Invitrogen).

Techniques: Activation Assay, Expressing, In Vitro, Transfection, Quantitative RT-PCR, Control, Immunocytochemistry, Staining, Western Blot, Microscopy

Journal: Phytomedicine : international journal of phytotherapy and phytopharmacology

Article Title: Magnolol regulates miR-200c-3p to inhibit epithelial-mesenchymal transition and retinoblastoma progression by modulating the ZEB1/E-cadherin axis in vitro and in vivo.

doi: 10.1016/j.phymed.2022.154597

Figure Lengend Snippet: Fig. 8. ZEB1 mediates Wnt16/β-catenin/ZEB1 pathway activation and EMT in vitro. Y79 cells were transfected with ZEB1-siRNA for 48 h. (a) mRNA expression of Wnt16, β-catenin, ZEB1, E-cadherin, FN1, α-SMA, and VEGFA was examined through RT-qPCR. Ns, no significance, *p < 0.05, **p < 0.01, ****p < 0.0001 compared with the control group. (b–c) Protein expression of ZEB1 in Wnt16, E-cadherin, β-catenin, ZEB1, FN1, α-SMA, and VEGFA was examined through immunocyto chemistry staining and western blotting. Scale bar of confocal microscope image = 4 µm. Cytosolic (CP) and nuclear protein (NP). Each group: n = 3.

Article Snippet: Briefly, the cells were cultured on glass coverslips, washed with cold phosphate-buffered saline (PBS), and fixed with 4% paraformaldehyde in PBS at 4 ◦C for 15 min. After blocking, the cells were incubated with primary antibodies against Wnt16 (Santa Cruz), Y.-H. Lai et al. Phytomedicine 110 (2023) 154597 β-catenin (Gene Tex), ZEB1 (Novusbio), E-cadherin (Cell Signaling Technology), FN1 (Gene Tex), and VEGFA (Santa Cruz) overnight at 4 ◦C, rinsed with PBS, and incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature, followed by staining with 4’,6-diamidino-2-phenylindole (DAPI; Invitrogen).

Techniques: Activation Assay, In Vitro, Transfection, Expressing, Quantitative RT-PCR, Control, Staining, Western Blot, Microscopy

Journal: Phytomedicine : international journal of phytotherapy and phytopharmacology

Article Title: Magnolol regulates miR-200c-3p to inhibit epithelial-mesenchymal transition and retinoblastoma progression by modulating the ZEB1/E-cadherin axis in vitro and in vivo.

doi: 10.1016/j.phymed.2022.154597

Figure Lengend Snippet: Fig. 9. Schematic representation the putative mechanisms underlying MAG-mediated increase in antitumor miRNA miR-200c-3p expression to suppress EMT signaling pathway and Wnt16/β-catenin/ZEB1 axis activation in retinoblastoma.

Article Snippet: Briefly, the cells were cultured on glass coverslips, washed with cold phosphate-buffered saline (PBS), and fixed with 4% paraformaldehyde in PBS at 4 ◦C for 15 min. After blocking, the cells were incubated with primary antibodies against Wnt16 (Santa Cruz), Y.-H. Lai et al. Phytomedicine 110 (2023) 154597 β-catenin (Gene Tex), ZEB1 (Novusbio), E-cadherin (Cell Signaling Technology), FN1 (Gene Tex), and VEGFA (Santa Cruz) overnight at 4 ◦C, rinsed with PBS, and incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature, followed by staining with 4’,6-diamidino-2-phenylindole (DAPI; Invitrogen).

Techniques: Expressing, Activation Assay